Process for sterilizing foodstuffs and beverages

ABSTRACT

This invention relates to a process for sterilizing foodstuffs and beverages contaminated with spore-bearing microorganisms by adding thereto an enzyme selected from the group consisting of cellulase and lysozyme and thereafter heating the foodstuff or beverage to effect relatively low temperature sterilization.

United States Patent [191 Nagano et al.

[ 1 Dec. 3, 1974 PROCESS FOR STERILIZING FOODSTUFFS AND BEVERAGES [76] Inventors: Yuji Nagano, 9-13, 3-Chome,

Tsurukawa, Machida-shi; Iwao Sato, 15-51, 3 Chome, Asahi-machi, Machida-shi, both of Tokyo, Japan [22] Filed: July 27, 1973 [21] App]. No.: 383,433

[30] Foreign Application Priority Data UNITED STATES PATENTS 3,672,957 6/1972 Scharpf 195/96 OTHER PUBLICATIONS Chemical Abstracts, Vol 72, 1970, 52114w, Spores, Gould et al., (1968) 4, 276286.

Primary Examiner-Raymond N. Jones Assistant ExaminerEsther L. Massung Attorney, Agent, or FirmWo|der & Gross 5 7 ABSTRACT This invention relates to a process for sterilizing foodstuffs and beverages contaminated with spore-bearing microorganisms by adding thereto an enzyme selected from the group consisting of cellulase and lysozyme and thereafter heating the foodstuff or beverage to effect relatively low temperature sterilization.

5 Claims, No Drawings BACKGROUND OF THE INVENTION In general, sterilization by heating is most commonly applied for preparation of foodstuffs and beverages to be stored for a long period of time. For example, in heat sterilization of canned foodstuffs or beverages, putrefying microorganisms are completely sterilized by heating at a high temperature, e.g., 120C. The sterilized foodstuff or beverage can then be stored for a long period of time. However, due to heating at such high temperatures the taste and qualities of the foodstuff or beverage may deteriorate.

On the other hand, heat sterilization at a relatively low temperature, e.g., 7090C has been applied to sterilization of pouched foodstuffs and beverages. In such low temperature sterilization, the freshness of the foodstuff and beverage is retained over an extended period of time. However, because putrefying microorganisms are not completely sterilized, the foodstuff and beverage cannot be stored for a long period of time. Putrefying microorganisms which remains alive after heating at a low temperature are spore-bearing microorganisms.

It is known that heat resistance or spore-bearing microorganism before germination is higher than that after germination. In order to completely sterilize the microorganism before germination, it is necessary to heat the microorganism at a high temperature. However, as described hereinbefore, the taste and quality of foodstuffs and beverages which are heated at a high temperature deteriorate.

DETAILED DESCRIPTION OF THE INVENTION It has now been discovered that the improved sterilization of foodstuffs and beverages contaminated with spore-bearing microorganisms can be achieved by adding to the foodstuff or beverage (1) an enzyme capable of accelerating the germination of said microorganism, or (2) an enzyme capable of injuring the spores of said microorganism so as to reduce its heat resistance and thereafter heating the foodstuff or beverage.

The foodstuff or beverage can then be sterilized at relatively lower temperatures, e.g., less than 100C., preferably 75-95C and sterilization of the. foodstuff and beverage without loss of quality, taste or deterioration of freshness is thereby realized. Typical sterilization periods, such as to 60 minutes, preferably 10 to minutes are used.

As the enzyme capable of accelerating the germina-.

organism loses its heat resistance. This phenomenon is due to the germination of spores by the enzymatic action according to the present invention.

As the enzyme capable of injuring the spores of spore-bearing microorganism, for example, lysozyme (which is present in animal sources such as egg white) may be used. When spore-bearing microorganism contacts lysozyme before germination, its heat resistance (which can be determined, e.g. by D-value, the time required to destroy 90% of the organism see Modern Food Microbiology, by J. M. .la'y, pages 158, 195) significantly reduces, and subsequent heat sterilization can be easily effected at a lower temperature. The reduction of heat resistance is caused by the partial injury of the spore walls by the enzymatic action but the spore is not yet killed.

According to the process of the present invention, heat sterilization at a low temperature is effected by either accelerating the germination of spore-bearing microorganism or injuring the spore walls of the microorganism whereby heat resistance of the microorganism is reduced. Both the germination and injury of spores occur even when the treatment of enzymes are carried out in water or a buffer solution without nutrients. The enzymatic reaction easily proceeds in foodstuff or beverage containing nutrients.

The treatment with enzyme according to the present invention is usually carried out at a temperature of from 35C to C at a pH offrom 3.5 to 8.0. Normally, the amount of enzyme will be 0.05% to 10% preferably 0.1% to 0.5% based on weight of foodstuff. Typical treatment are 30 minutes to 120 minutes preferably to minutes. However, the conditions for the enzymatic treatment such as pH, temperature, concentration of enzyme, etc., vary, depending upon the foodstuff or beverage to be treated, putrefying microorganism and the enzyme in use. It is therefore necessary to select suitable conditions to the foodstuff or beverage to be treated.

The following examples 1 and 2 exemplify the sterilization effect on the enzymes on some typical sporebearing microorganisms.

All ATCC members refer to microorganisms on deposit with the American Type Culture Collection and are fully available to the public.

Example 1 Spores of Bacillus subtilis ATCC 6633 were suspended in a phosphate buffer solution which was then subjected to filtration to prepare a spore suspension containing 10 spores per ml. 10 ml. of 0.5% solution of Driselase (trade name for a microbiological enzyme composition consisting of cellulase, glucanase, xylanase, pectinase, dextranase, amylase, protease and substances having macerating activity, available from Kyowa Hakko Kogyo Kabushiki Kaisha, Japan) was added to the spore suspension (20 ml.) obtained. Thereafter, the enzymatic reaction was effected at 37C for 60 minutes.

The germinating rate of the spores in the reaction mixture was determined periodically by microscopic observation using the methylene blue staining method. The results obtained are shown in the following table, from which it is confirmed that about 99% of all spores germinated for 60 minutes. Each reaction mixture periodically determined was heated at C for 10 minutes and then the number of the living cells in the reaction mixture was counted. As apparent from the table, the greater the increase of germinating rate, the greater the decrease of the living cells after heating. After treatment, with enzyme for 60 minutes, sterilization is effected at 95C for 10 minutes.

TABLE A' Time of enzyme Germinating Number of living cells As shown in Table B, the sterilization of soup was effected at l20C with or without the-addition of enzyme. When the soup was not treated with enzyme," ther'e were living cells therein after heating at a temperature below 90C, and the soup putrefied during preservation at a room temperature for 2 or 3 days.

On the contrary, when the soup was treated with Driselase pursuant to the present invention (Sample B) treatment (mm') me (cells after heamig) 5 and sterilized by heating at 90C, it did not putrefy even I 0 0 5.4 x after preserving for more than 6 months. Sample B 10 l8 heated at 90C had better taste when compared with 2g 3g 2 8 8 l0 Sample A heated at l20C. 9O 99 Example 4 10 By using pork meat (8 kg), beef meat (4 kg.) and In the case of the spores of Bacillus subtilis which are on meat g-) sausages were p p in a resistant to heat, it is normally necessary to heat the Ventlonal y Curing, mlncing, kneafllng spores at 120C for 20 minutes for sterilization." Howgethe? stuffmgo mto Parts F intestine and ever, since according to the present invention the Smoking at 80 C for 40 i boiling m a water for spores germinate and consequently lose their resistance minutes then Cooling to heat when treated with Driselase sterilization may If] the kneading Stage an aqueous SOhm-on comalm-ng o i Driselase was added to the minced meats in the amount now be effected at 95 C for 10 minutes. Such sterihza- Shown in the following condmonsi w 95 C, p pl'eserve q l ty n The sausages obtained were left standing by hanging freshness of the feodstuff beveragei Y a 20 in acontainer adjusted to a temperature of 30C and a Example 2 relative humidity of 90% for a period as shown in the following table, to determine the degree of putrefactlon Spores of Closlridium thermosaccharolyticum were" of the sausagessuspended in an ordinary medium having a pHof 7.0' and containing yeast extract (2.5 g) polypeptone (5.0g) and glucose (l-.() g) per liter of-mediunn Egg' A oumof standing period (day) white lysozyme was added to the medium in aconcenadded enzyme 2 3 .4 5 tration of up to 0.05% by weight of the medium. After Y 0% (none) slime -H-++ this, the mixture was subjected to reaction at 37C-for 0.5% gag? I f 1* 1: 1 60 minutes, and then to heating at 90C for lOminutes. putrid smell i No living cells are observed in the heated reaction mixm fjggf sn'm" I I I I I ture.

Neither-germination nor death of spores-wasob ifjj meat served at the completion stage of the enzymatic treate ment. It was confirmed that the spores of Clostridium thermosaccharolyticum were partially injured by the lye; A5 appareni fmm.the above the addiuon of sozyme treatment so as to reduce heat resistance, and genulasednakes posslble 5 im Sausages 'g v The followmg examples further As described above, according to the present inventhe present mvennon' 40 tion, it is possible to completely sterilize foodstuffs and Example 3 beverages contaminated with spore-bearing microorgamsms at a lower temperature by germinating the T0 p g P was added Driselase y spores or partially injuring the spores so as to reduce weight) and the composition then well mixed. Samples their heat resistance, and permit effective sterilization of potage soup (each. 100 g) were separately placed in at low temperatures. pouches d f a lli d l i fil d h That whichis sought to be protected is set forth in the pouches sealed in vaccuo. After standing for'30 minfollowmg clalmsutes, at a temperature at 30C samples were respecclam: tively heated at l20C, 90C and 70C for'20 minutes. A Process for stflllzms foodstuffs and beverages by m cans of Autoclava ctt ntemilng spore-beatrmg nt icrotorglamstms by tusel of rela we y ow empera ures or s eriiza 1011 W [C comfollovi'mg. table Shows the number of i which prises adding thereto an enzyme selected from the remained alive in the samples after enzymatic and heat group consisting of (a) cellulase, and (b) lysozyme for treatmentsa treatment period, and then subjecting said foodstuff TABLE 3 i 5 5 and beverage to relatively low temperature sterilization conditions. Number of 2.'The process of claim 1 wherein said cellulase in the Heating temperature No addition Addition or f form of an enzyme preparate further containing minor after of ym Driselzise amounts of glucanase, xylanase, pectinase, dextranase, Sample enzyme treatment (celllmil) (cell/mil.) amyiase and protease A [20C 0 0 3. The process of claim 1 wherein a sterilization tem- B 90C I 4 10 v0' pe'rature of less than 100C is used. c C I 2 x l0 3.3x i of 4, The process of claim 1 wherein said treatment is normally from 30 to 120 minutes at 35C to 55C with a pH of 3.5 to 8.0.

5. The process of claim 1 wherein said sterilization step is conducted at temperatures of C to C for 10 to 60 minutes. 

1. A PROCESS FOR STERILIZING FOODSTUFFS AND BEVERAGES CONTAINING SPORE-BEARING MICROORGANISMS BY USE OF RELATIVELY LOW TEMPERATURES FOR STERILIIZATION WHICH COMPRISES ADDING THERETO AN ENZYME SELECTED FROM THE GROUP CONSISTING OF (A) CELLULASE, AND (B) LYSOZYME, FOR A TREATMENT PERIOD, AND THEN SUBJECTING SAID FOODSTUFF AND BEVERAGE TO RELATIVELY LOW TEMPERATURE STERILIZATION CONDITIONS.
 2. The process of claim 1 wherein said cellulase in the form of an enzyme preparate further containing minor amounts of glucanase, xylanase, pectinase, dextranase, amylase and protease.
 3. The process of claim 1 wherein a sterilization temperature of less than 100*C is used.
 4. The process of claim 1 wherein said treatment is normally from 30 to 120 minutes at 35*C to 55*C with a pH of 3.5 to 8.0.
 5. The process of claim 1 wherein said sterilization step is conducted at temperatures of 75*C to 95*C for 10 to 60 minutes. 